PROJECT SUMMARY Congenital cardiac anomalies are the most prevalent birth defects, affecting over 1% of live births. Despite remarkable progress in understanding cardiac development, the mechanisms underlying cardiac maldevelopment in embryos that result in malformations are largely unknown. Numerous genetic mutations resulting in cardiac maldevelopment have been identified, however the wide range of symptoms and severity of these mutations suggest a prominent role of epigenetic (non-genetic) aggravating factors in the development of cardiac anomalies. Our overarching hypothesis is that both genetic and epigenetic factors implicated in the development of cardiac anomalies alter the biophysical properties of cardiac tissue and that these properties dynamically regulate developing cardiomyocytes and non-myocytes in vitro. This proposal seeks to analyze the biophysical properties of embryonic cardiac tissue, such as mechanical properties and extracellular matrix (ECM) composition, and use this information to build 3D hydrogel tissue mimics of normal and diseased cardiac tissues. In Specific Aim 1, we will engineer ECM-based scaffolds to mimic and study the biophysical properties of cardiac tissue with genetically induced anomalies. In Specific Aim 2 we will engineer ECM-based scaffolds to mimic and study the biophysical properties of cardiac tissue with epigenetically induced anomalies. The 3D hydrogel cardiac tissue mimics developed in each aim will be used to investigate the interplay between biophysical properties and cell phenotype/genotype in the development of cardiac anomalies. Development of 3D cardiac tissue mimics has the potential to elucidate both pathological mechanisms of disease and the biophysical properties that result in physiologic cellular development for engineering cardiac tissues. Information obtained in this project will identify potential ECM targets for genetic therapy, and future studies will utilize our cardiac tissue mimics as a base for development of an injectable biomaterial to treat cardiac anomalies in humans.